Trait and edaphic variable measurements
The green leaf surface areas were
measured by scanning at least five fully expanded leaves using a
portable leaf-area meter (Li-3000C, LI-COR, USA), and their area was
determined using ImageJ software (NIH Image, Bethesda, USA). For
gymnosperm species, 20 leaves per
tree were scanned to ensure accuracy. The foliage samples were
oven-dried (60 °C, 48 h) and weighed to determine the dry mass. Specific
leaf area was expressed as the ratio of leaf area to leaf dry mass.
Leaf
thickness was measured using digital callipers (SMCTW Company, Shanghai,
China) to avoid major veins. Leaf tissue density was then calculated as
dry mass divided by the product of leaf area and leaf thickness
(Fortunel et al. 2012).
The first- and second-order roots of all species were selected as
absorptive roots, because they are considered as belowground
resource-acquiring units (Xia et al. 2010; McCormack et al. 2015). To
obtain the morphological traits accurately, the dissected root samples
were cleaned with deionised water, spread out on glass panes to avoid
overlapping, and scanned using an Epson Expression 10000 XL scanner
(Seiko Epson Corporation, Japan) at a resolution of 400 dpi. Root
diameters and ARL were measured using
WINRHIZO version 2012b (Regent
Instruments Inc., Quebec City, Quebec, Canada). Subsequently, the
scanned samples were oven-dried (60 °C, 48 h) and weighed to calculate
SRL (root length divided by its dry mass) and RTD (root dry mass divided
by its volume; assuming the root to be a cylinder). We calculated BI as
the ratio of the 1st- and 2nd-order root number to the 3rd order root
length (Kong et al. 2014).
The oven-dried samples of green leaves, senesced leaves, and absorptive
roots were ground to fine powder using a Retsch MM 400 mixer mill
(Retsch GmbH, Haan, Germany) for chemical analyses. The concentrations
of C and N were determined using a Vario MACRO cube elemental analyser
(Elementar Analysensysteme GmbH, Langenselbold, Germany). The P
concentration was determined using an inductively coupled plasma mass
spectrometer (Optima 5300 DV, Perkin Elmer, Waltham, USA). The lignin
concentration in green leaves was analysed using the widely used acid
detergent lignin method (van Soest & Wine 1968). Ash contents were
determined by combusting 40–50 mg of the corresponding samples in a
muffle furnace (550°C for 4 h). All initial chemical traits and residual
mass of decomposing leaves (see below) were expressed on an ash-free,
dry-weight basis.
Soil nitrate (NO3-)-N and ammonium
(NH4+)-N were extracted from the soils
(2 mol l-1 KCl, 50 ml), and their concentrations were
determined using a continuous-flow autoanalyser (Autoanalyzer 3, Bran
and Luebbe, Germany). Available P (AP) was extracted with
KCl/NH4F, and its concentration was determined
colourimetrically using ascorbic acid molybdate analysis on a
continuous-flow autoanalyser.